Members of the genus Chlamydia are obligate intracellular parasitic bacteria and the agents of various animal and human diseases. Infections caused by C. trachomatis are now recognized as the most prevalent - and among the most damaging - of all sexually transmitted diseases seen in the United States. Chlamydiae possess a unique life cycle which may be thought of as the orderly alternation between infectious extracellular elementary bodies (EBs) and noninfectious intracellulary dividing reticulate bodies (RBs). Almost nothing is known about the factors which govern EB entry into host cells, reorganization of EBs to RBs, multiplication of RBs, and reorganization of RBs to EBs. The overall objective of this proposal is to better understand the molecular events which take place during the developmental cycle of C. trachomatis. The long term goal is to establish conditions for host-free growth of chlamydiae, an achievement which will greatly facilitate basic studies on these difficult-to-grow organisms. We will identify genes expressed at specific phases of the developmental cycle and establish a library of these differentially expressed genes. Phase-specific protein synthesis and the DNA library will be used to monitor gene expression in host-free chlamydiae incubating under a variety of conditions in order to identify environmental signals which control chlamydial reorganization and promote multiplication. Surface proteins synthesized at the time of RB to EB conversion will be identified and purified, and their roles as adhesins will be examined. A recombinant drug-resistant plasmid will be constructed, and attempts will be made to introduce this plasmid into C. trachomatis in order to develop a system for studying the genetics of chlamydiae. The identification of genes involved in the reorganization of chlamydial life-cycle forms and the determination of environmental signals which control their expression should ultimately improve our understanding of how chlamydiae survive within the human host and cause persistent infections. Identification of a chlamydial adhesin might provide the basis for the development of a subunit vaccine.